A new inhibitor of phospholipase A₂ was isolated from the fermentation broth of Aspergillus unguis. The structure, with a depsidone carbon skeleton, was assigned by spectroscopic experiments.

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Gliocladium sp. FO-1513 was found to produce novel inhibitors of acyl-CoA: cholesterol acyltransferase (ACAT). Two active compounds, designated glisoprenins A and B, were isolated from the culture broth of the producing strain by a conventional method. The IC₅₀ values of glisoprenins A and B for ACAT activity were 46 and 61 JUM in an enzyme assay using rat liver microsomes, and 1.2 and 0.57μM in a J774 macrophage assay, respectively.

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New cyclodepsipeptides named enniatins D, E and F were isolated from the culture broth of Fusarium sp. FO-1305 as inhibitors of acyl-CoA: cholesterol acyltransferase (ACAT). The respective structures of enniatins D, E and F were determined to be cyclo[D-α-hydroxyisovaleryl(D-Hiv)-L-N-methylleucinyl (L-Me-Leu)-D-Hiv-L-N-methylvalinyl(L-Me-Val)-D-Hiv-L-Me-Val], a mixture of cyclo[D-Hiv-L-Me-Leu-D-Hiv-L-N-methylisoleuciny (L-Me-He-D)-D-Hiv-L-Me-Val], and cyclo(D-Hiv-L-Me-Ile-D-Hiv-L-Me-Leu-o-Hiv-L-Me-Val), and cyclo (D-Hiv-L-Me-Leu-D-Hiv-L-Me-Ile-D-Hiv-L-Me-Ile) by spectral analyses and chemical degradation. The IC₅₀ values of enniatins D, E and F for ACAT activity in an enzyme assay using rat liver microsomes were calculated to be 87, 57 and 40μM, respectively.

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Acaterin, a novel inhibitor of acyl-CoA: cholesterol acyltransferase (ACAT), was isolated from a culture broth of Pseudomonas sp. A92 by Diaion HP-20 column chromatography, solvent extraction and reverse phase HPLC. Spectroscopic analyses of the compound yielded 3-(1-hydroxyoctyl)-5-methyl-(5H)-furanone as the proposed structure. In the presence of oxidized low density lipoprotein, acaterin inhibited the synthesis of cholesteryl ester in macrophage J774 by 50% at a concentration of 45 μM. Acaterin also inhibited AC AT activity in the rat liver microsomes by 50% at a concentration of 120μM. Kinetic studies suggested that inhibition of ACAT by acaterin was noncompetitive with respect to oleoyl-CoA.

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A10255 is a complex of new thiopeptide antibiotics characterized structurally by a cyclic peptide core to which is attached a side chain composed of dehydroalanine moieties. The complex contained 80-85% factor B, 15-20% factor G, and trace amounts of factors C, D, E, F, H, and J. Taxonomic studies indicated the producing microorganism to be a strain of Streptomyces gardneri. The major portion of the antibiotic produced remained associated with the mycelial biomass, from which it was extracted with polar solvents such as aqueous methanol or aqueous acetone. Initial A10255 yields of <2μg/ml were increased to over 300μg/ml in stirred reactors through strain selection, nutritional studies, and conversion of the batch fermentation to a fed-batch mode.

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Angolamycin (1) and two novel analogues (2 and 3) were isolated from the culture broth of a Streptomyces strain. NMR and MS analysis proved that 2 is the 18-dihydro-, while 3 is the 18-deoxo-18-dihydro derivative of angolamycin. Full experimental assignment of the ¹H and the ¹³C NMR spectra of these compounds was obtained from ID and 2D chemical shift correlation measurements. Compounds 2 and 3 are less potent antibiotics than angolamycin.

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Streptomyces roseiscleroticus L827-7 (ATCC 53903) produced a novel antifungal and antitumor antibiotic, sultriecin. It exhibited in vitro antifungal activity and potent in vivo antitumor activity against P388 and L1210 leukemias, and B16 melanoma. Sultriecin is composed of several unique structural units; a conjugated triene, an α, β-unsaturated (5-lactone, and a sulfate functionality.

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Kedarcidin, a new chromoprotein antitumor antibiotic, was isolated from the culture broth of a novel actinomycete strain L585-6 (ATCC 53650). The antibiotic was recovered from the culture nitrate by adsorption to QAE ion exchanger and purified by successive application of gel filtration and ion exchange chromatography with Sephadex G-50 and DEAE-Sephadex, respectively. Kedarcidin is an acidic complex (pI 3.65) with an apparent molecular weight of 12, 400. The complex consists of a highly unstable, solvent extractable chromophore and a water soluble peptide. The apoprotein is a single chain polypeptide of 114 residues.

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Biosynthetically blocked mutants were obtained from a baumycin-producing Streptomyces sp. D788 newly isolated from soil. The first mutant isolated was a baumycin-negative but daunorubicin-accumulating mutant with a loss of 4'-substitution activity, from which all other blocked mutants were successively derived. These included a known 11-deoxydaunorubicin-producing mutant and several new types of mutants which produced mainly 10-carboxy-13-deoxocarminomycin, 10-methoxycarbonyl-13-deoxocarminomycin, their 11-deoxy derivatives or a precursor aglycone, respectively.
In this paper, all the anthracycline components produced by the parent strain and its two known blocked mutants, a daunorubicin producer and a 11-deoxydaunorubicin producer, are also determined by HPLC and five new components are isolated. Cytotoxicities in vitro of all the components against L1210 cell culture are also described.

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Vermixocins A and B, 3-(1'-hydroxy-3'-methylbutyl)- and 3-(1'-acetoxy-3'-methylbutyl)-11-hydroxy-4-methoxy-9-methyl-5H, 7H-dibenzo[c, f][1, 5]dioxocin-5-one, respectively, were isolated from the mycelium of Penicillium vermiculatum. Both metabolites showed cytotoxic effects on lympholeukemia cells P388.

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Enniatin synthetases from the cyclodepsipeptide producers Fusarium lateritium and Fusarium sambucinum were purified to homogeneity and characterized. Like the previously described enniatin synthetase from Fusarium scirpi both enzymes consist of a single polypeptide chain and are very similar concerning their Mr (250 kdaltons) and reaction mechanism. Limited proteolytic digests show only slight differences in their patterns in SDS-gels. Interestingly the synthetases differ in their amino acids specificities. The enzyme from the enniatin A producer F. sambucinum exhibits a high affinity to the substrate amino acids L-Leu and L-Ile. In contrast the synthetase from the enniatin B producer F. lateritium preferably accepts L-Val, the constituent amino acid of enniatin B.

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A10255 is a complex of new thiopeptide antibiotics produced by Streptomyces gardneri. When stirred reactors were operated in batch mode using a denned medium with a glucose feed, 250 μg/ml of A10255 were produced during a four-day fermentation cycle. The linear growth phase of S. gardneri was extended through seven days by supplementing the defined medium with continuous feeds of hydrolyzed casein and methyl caprate. With the supplementary feeds, antibiotic biosynthesis paralleled growth during the extended cycle and attained levels of 1, 750 μg/ml. Increasing the standard glucose feed rate increased titers principally by increasing cell mass. Supplementing the standard glucose feed with lipids such as caprylate or caprate, and decyl alcohol, affected cell mass minimally but produced higher titers by increasing the specific biosynthesis of A10255 per unit of biomass.

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Aklavinone is an aglycone of aclacinomycin A which is an important antitumor drug. Genes for the biosynthesis of aklavinone were cloned from Streptomyces galilaeus 3AR-33, an aklavinoneproducing mutant, by use of the actI and actIII polyketide synthase gene probes.Restriction mapping and Southern analysis of the DNA cloned in a λ phage vector established that the DNA represented three different regions of the S. galilaeus 3AR-33 genome that contained 3.4, 2.5, and 4.1kb BamHI fragments which hybridized with actIII. Of those, only the 3.4kb fragment also hybridized with actl. Complementation experiments with specifically blocked mutants confirmed that the cloned 3.4kb BamHI fragment contains the genes required for the early stage of polyketide synthesis in aklavinone biosynthesis.

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An antibiotic, prodigiosin 25-C, preferentially suppresses cytotoxic T lymphocytes (CTL) without affecting antibody production. Here, we investigated the effect of prodigiosin 25-C on delayed-type hypersensitivity (DTH), graft versus host reaction (GvHR) and allogeneic skin graft rejection. DTH reactions were markedly inhibited by ip treatment of the mice with prodigiosin 25-C. Cell transfer experiments indicated that prodigiosin 25-C exerted its suppressive effect on the late efferent phase rather than on the induction phase of DTH. Prodigiosin 25-C suppressed induction of anti-host CTL when GvHR was induced by iv inoculating splenocytes of parental C57BL/6 mice to adult unirradiated BDF₁ mice. It had little effect on GvHR-induced splenomegaly observed 2 weeks after the inoculation, but significantly delayed the subsidence of splenomegaly as revealed 8 weeks later, suggesting that suppression of CTL converts immunosuppressive GvHR to immunostimulative one as reported by G. M. SHEARER. However, reduction of interleukin-2 (IL-2) production and mitogen responses induced by GvHR were not rescued by prodigiosin 25-C treatment. Prodigiosin 25-C moderately prolonged survival of major histocompatibility (MHC)-mismatched skin grafts. Since the mode of action of prodigiosin 25-C is distinct from those of cyclosporin A and FK506, these results demonstrate potential usefulness of the antibiotic for a supplementary immunosuppressant.

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Proliferation of concanavalin A (Con A)-activated nylon-wool purified murine splenic T cells was increasingly suppressed by prodigiosin 25-C as higher concentrations of Con A were used for the activation. Enhancement of suppressive effect of prodigiosin 25-C was not observed when T cells were stimulated with phytohemagglutinin (PHA), anti-CD3 antibody, or allogeneic splenic adherent cells. The suppressive effect of prodigiosin 25-C was enhanced by the addition of Con A in various T cell subpopulations as well as in LPS-activated splenic B cells. Lectins that recognize mannose residue of biantennary-complex-type sugar chains significantly enhanced the suppressive effect of prodigiosin 25-C, whereas a lectin that binds to N-acetylglucosamine did not. These results suggest that binding of lectins to the mannose residue of biantennary-complex-type sugar chains on cell surface of both T and B lymphocytes plays a central role on the enhancement of the suppressive effect of prodigiosin 25-C.

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In an attempt to improve the isolation of the antibiotic rubradirin from fermentations of Streptomyces achromogenes var. rubradiris, the use of preparative reversed-phase chromatography was investigated. The product isolated was a mixture of rubradirin and a new antibiotic named protorubradirin, of extremely similar structure, which is converted into rubradirin on exposure to light and air. Methanolysis of protorubradirin in the dark yields an anomeric mixture of methyl glycosides of a C-nitroso-sugar, converted photo-oxidatively into the methyl rubranitrosides derived from rubradirin. Thus, protorubradirin is the C-nitroso-analogue of rubradirin. It is suggested that the same relationship between protorubradirin and rubradirin may apply to the anthracycline antibiotics viriplanin A and viriplanin D.

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The structural characterization of minor components of bacitracin (BC) complex was carried out using a technique of liquid chromatography/mass spectrometry (LC/MS). Satisfactory total ion current chromatogram of BC complex and excellent mass spectra of many components were given by Frit-fast atom bombardment (FAB) LC/MS analytical system, and the structures of 13 minor components could be proposed. The 13 minor components were classified into two groups, bacitracin A (BC-A) related components and bacitracin F (BC-F) related components depending on their common N-terminal moieties. The structures of BC-A related components and BC-F related components were the same as those of BC-A and BC-F, respectively, except that one to three of isoleucine and leucine residues are replaced by valines. The BC-F related components were degradation products of BC-A related components through the same degradation process as that of BC-A.

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The bactericidal activity of cefclidin (E1040), a new cephalosporin, against a clinical strain of Citrobacter freundii was compared with that of ceftazidime in a two-compartment in vitro pharmacokinetic model system designed to simulate plasma concentrations in humans for 12 hours after intravenous administration of a 1 g dose. Both cefclidin and ceftazidime showed rapid bactericidal activity against C. freundii. However, during the simulation of ceftazidime treatment, regrowth was observed after two hours and a subpopulation emerged which was resistant to ceftazidime. Neither regrowth nor the emergence of resistant mutants was observed with cefclidin during the 12-hour simulation. The ceftazidime-resistant mutants constitutively overproduced β-lactamase at levels which were about 500-fold higher than that of the parent wild-type strain. Against this β-lactamase overproducing mutant, no bactericidal activity of ceftazidime was observed in the in vitro model system, whereas the bactericidal activity of cefclidin was observed during the 12-hour period. The emergence of Enterobacter cloacae mutants derepressed for β-lactamase production was also observed with ceftazidime but not cefclidin. The affinity of cefclidin for the β-lactamase isolated from these mutants was lower than that of ceftazidime, and the kinetic parameters of enzymatic hydrolysis showed that cefclidin was hydrolyzed more slowly at a low concentration (0.2μM) than was ceftazidime. It is suggested that the high activity of cefclidin against strains derepressed for β-lactamase plays a major role in the absence of emergence of resistant mutants.

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The binding of some cephalosporins to human serum albumin (HSA) was studied by an ultrafiltration technique. Changes in C-3 side chain resulted in marked changes in the binding to HSA, but changes in C-7 side chain did not. Cephalosporins were classified into three groups by C-3 side chain: (i) Cationic side chain with low affinity for HSA; (ii) anionic side chain with high affinity for HSA; (iii) non ionized side chain, in which binding to HSA was dependent on lipophilicity. These findings suggest that electrostatic and hydrophobic forces play a role in the binding affinity of cephalosporins for HSA. The binding of cephalosporins with high HSA affinity was displaced significantly by warfarin but not by phenylbutazone, L-tryptophan, or diazepam. The interaction of the cephalosporins with high affinity for HSA with chemically modified HSA was investigated to clarify the amino acid residues of HSA involved in the cephalosporin binding sites. The binding of the cephalosporins decreased remarkably with the modification of the tyrosine residues. These results suggest that the binding site of cephalosporins is located in the vicinity of warfarin binding site rather than benzodiazepine binding site and that tyrosine residues are involved in the cephalosporin binding site.

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2-(Alkyloxycarbonyl)-2-alkylideneethyl esters of various aminothiazole-oxyimino cephalosporins have been synthesized and studied. They are useful alternatives to the currently existing orally active esters. Among the new esters synthesized, the 3'-azidomethyl cephem ester Ro 41-3399 (7k) presented an oral bioavailability superior to the corresponding pivaloyloxymethyl ester (9) in a rat model and was selected as a candidate for further evaluation.

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